Being one tissue in the mammalian body that regenerates in postfetel life, bone is a promissing tissue for research on morphogenetic matrix for osteogenetic gene expression. The proposed research deals with the follwoing hypothetical constituents: the bone morphogenetic property (BMP), an insoluble polypeptide firmly attached to collagens of bone, dentin, and other calcified tissues; neutral proteinase (BMPase), geographically in close association with, but separable from BMP; BMPase-inhibitor, a soluble non-collagenous protein surrounding BMPase; BMP-inhibitor, a high molecular weight soluble protein in juxtaposition to BMP. The investigative procedures are designed to separate and purify the constituents of the bone morphogenetic system either by digestion of bone matrix with collagenase, or by chemical extraction without enzymic degradation or by the combination of the two. The methods include: filtration of digest or extract on gel columns at different levels of pH; chromatography on ion-exchangers; paper or column electrophoresis under acidic conditions. The morphogenetic constituents are demonstrable and bioassayed in: (a) residues of matrix implanted in muscle in rats; (b) isolated substances added to chemically-defined culture media in vitro in systems consisting of mesenchymal cell outgrowths of muscle into substrata prepared from denatured bone matrix; (c) isolated proteins implanted in double and triple walled millipore and nuclepore chambers in muscle pouches with and without minced or disaggregated muscle mesenchymal cells. Bibliographic references: Urist, M.R.,Nogami, H. and Terashima, Y.: A substratum of bone gelatin for chondrogenesis in tissue culture and in vivo. In: Extracellular Matrix Influences in Gene Expression. H.C. Slavkin and R. Gruelich, eds. pp. 609-618, New York, N.Y., Academic Press, 1975. Urist, M.R., Mikulski, A. and Boyd, S.D.: A chemosterilized antigen-extracted bone morphogenetic alloimplant. Arch. Surg. 110:416-428, 1975.